Anat Cell Biol.  2023 Mar;56(1):122-136. 10.5115/acb.22.204.

Silymarin attenuates escitalopram (cipralex) induced pancreatic injury in adult male albino rats: a biochemical, histological, and immunohistochemical approach

• Salama RM
• Tayel SG

• KMID: 2540991
• DOI: http://doi.org/10.5115/acb.22.204

Abstract

Depression is a prevalent global problem since ages, predominately treated with SSRI. Cipralex, is an antidepressant of the SSRIs class used as a remedy for mood, depression and anxiety. Silymarin (SIL), a natural free radical scavenging, has an antioxidant and anti-inflammatory properties. This hypothesis evaluates, for the first time, the role of cipralex on the structure of the endocrine and exocrine components of the pancreas and assess the beneficial effects of SIL on these changes. Forty-five rats were divided into control, cipralex, and cipralex plus SIL groups. During sacrifice, all rats and pancreases were weighed and the ratio of pancreatic weight (PW) to rat weight (RW) was calculated, blood samples were collected to estimate fasting glucose, insulin and amylase levels, the specimens were prepared for histological, immunohistochemical (inducible nitric oxide synthase [iNOS], tumour necrosis factor-alpha [TNF-α], caspase 3, proliferating cell nuclear antigen [PCNA], and anti-insulin antibody), and morphometrical studies. Cipralex group exhibited marked destruction of the pancreatic architecture of the exocrine and endocrine parts, with a dense collagen fiber deposition. Also, there is highly significant decrease (P<0.001) of PW/RT ratio, insulin, and amylase levels, the number and diameter of islets of Langerhans, the number of PCNA positive immunoreactive cells, and the number of insulin positive β-cells. Furthermore, a highly significant increase of glucose level, iNOS, TNF-α, and caspase-3 positive immunoreactive cells in the islets of Langerhans and acinar cells were observed. SIL improves the pancreatic histological architecture, weight loss, biochemical, and immunohistochemical analyses. Administering SIL is advantageous in managing cipralex induced pancreatic injury via its anti-inflammatory, antioxidant, and anti-apoptotic qualities.

Keyword

Cipralex; Silymarin; Inducible nitric oxide synthase; Tumor necrosis factor-alpha; Proliferating cell nuclear antigen

Figure

• 

  Fig. 1 A histogram demonstrating a highly significant decrease (P<0.001) in the PW/BW ratio in the cipralex group compared to the control group, while the cipralex plus SIL group, illustrating a significant increase compared to the cipralex group, with no significant difference between control and cipralex plus SIL groups. PW, pancreatic weight; RW, rat weight; SIL, silymarin.

• 

  Fig. 2 A histogram illustrating a highly significant increase (P<0.001) in FBG in the cipralex group compared to the control group, while the cipralex plus SIL group, illustrating a high significant decrease compared to the cipralex group, with no significance between control and cipralex plus SIL groups. SIL, silymarin; FBG, fasting blood glucose.

• 

  Fig. 3 A histogram showing a highly significant decrease (P<0.001) in the insulin level in the cipralex group compared to the control group, while the cipralex plus SIL group, illustrating a significant increase compared to the cipralex group, with no significant difference between control and cipralex plus SIL groups. SIL, silymarin.

• 

  Fig. 4 A histogram demonstrating a highly significant decrease (P<0.001) in the amylase level in the cipralex group compared to the control group, while the cipralex plus SIL group, illustrating a high significant increase compared to the cipralex group, with no significance between control and cipralex plus SIL groups. SIL, silymarin.

• 

  Fig. 5 Representative H&E staining of rat pancreas; (A) Control group: demonstrating that the pancreas is formed of lobules (L) of different sizes and shapes separated by thin connective tissue septa (arrow). Interlobular duct (curved arrow) and blood vessel (BV) are observed. Each lobule is formed of highly packed darkly stained acini (A) (the exocrine portion of the pancreas). The islet of Langerhans (I) (the endocrine portion of the pancreas) is appeared as large defined pale stained area scattered between the darkly stained pancreatic acini. (B) Cipralex group: showing destruction of the normal pancreatic architecture of the exocrine part (E), congested interlobular BV and distorted fat cells (F) are observed. (C) Cipralex group also illustrating shrunken and vaguely identified pancreatic islet (I), distorted acini (A). Inflammatory cell infiltration (arrow) are also observed. (D) Cipralex plus SIL group: showing restoration of the normal architecture of the E and endocrine part (I). Interlobular BV and duct (arrow) appear normal in between the cells (×200, scale bar=40 μm).

• 

  Fig. 6 Representative H&E staining of rat pancreas; (A) Control group: illustrating that the pancreatic acini (A) have basal basophilic cytoplasm containing rounded vesicular nuclei (black arrow) and apical acidophilic zymogen granules (Z). The islet of Langerhans (I) is composed of cords of secretory cells separated by blood capillaries (curved arrow). Two types of cells are recognized; central beta cells with rounded and lighter nuclei (red arrow) and peripheral alpha cells with oval darkly stained nuclei (arrow head). (B) Cipralex group: showing that many acinar cells have darkly stained pyknotic nuclei (black arrow). Distorted shape of islet cells with apparent decrease in the number (I) and congested blood capillaries (arrow head) are noticed. Thickening in the wall of pancreatic duct (curved arrow) and thickening of the interlobular septa are observed (yellow arrow). (C) Cipralex group: also demonstrating that most of pancreatic acinar cells have cytoplasmic vacuolations (V) and many β-cells of islet of Langerhans are lost and marked vacuolations (red arrow) are seen. Empty spaces (*) are found between pancreatic acini. (D) Cipralex plus silymarin group: showing few acinar cells with pyknotic nuclei (black arrow) and some cells have mild vacuolated cytoplasm (v). Few β-cells appear vacuolated (red arrow) (×400, scale bar =20 μm).

• 

  Fig. 7 Representative Masson’s Trichrome stain of rat pancreas; (A) Control group: showing delicate collagen fibers around the pancreatic acini, pancreatic ducts and blood vessels (yellow arrow). (B) Cipralex group: showing dense collagen fiber deposition (yellow arrow). (C) Cipralex plus SIL group: showing moderate collagen fiber deposition (yellow arrow) as compared with cipralex group (yellow arrow) (×400, scale bar=20 μm). (D) A histogram demonstrating a highly significant increase in the mean area percentage of collagen fibers in cipralex group compared to control group. While cipralex plus SIL group showing a highly significant decrease compared to cipralex group, with no significant difference between control and cipralex plus SIL groups. SIL, silymarin.

• 

  Fig. 8 Representative iNOS immunostaining of rat pancreas; (A) Control group: illustrating negative immunoreactivity in pancreatic acini and islet cells. (B) Cipralex group: illustrating strong cytoplasmic immunoreactivity in most pancreatic acini and islet cells. (C) Cipralex plus SIL group: illustrating mild cytoplasmic immunoreactivity in pancreatic acini and islet cells as compared with cipralex group (×400, scale bar=20 μm). (D) A histogram illustrating a highly significant increase in iNOS immune expression in cipralex group compared to control group. While cipralex plus SIL group showing a highly significant decrease compared to cipralex group, with no significance between control and cipralex plus SIL groups. iNOS, inducible nitric oxide synthase; SIL, silymarin.

• 

  Fig. 9 Representative TNF-α immunostaining of rat pancreas; (A) Control group: revealing negative immunoreactivity in pancreatic acini and islet cells. (B) Cipralex group: showing strong immunoreactivity in most of pancreatic acini and islet cells. (C) Cipralex plus SIL group: showing mild immunoreactivity in pancreatic acini and islet cells as compared with cipralex group (×400, scale bar=20 μm). (D) A histogram showing a highly significant increase in TNF-α immune expression in cipralex group compared to control group. While cipralex plus SIL group showing a highly significant decrease compared to cipralex group, with no significant difference between control and cipralex plus SIL groups. TNF-α, tumour necrosis factor-alpha; SIL, silymarin.

• 

  Fig. 10 Representative caspase-3 immunostaining of rat pancreas; (A) Control group: demonstrating negative immunoreactivity in pancreatic acini and islet cells. (B) Cipralex group: demonstrating strong cytoplasmic and nuclear immunoreactivity for caspase-3 in most pancreatic acini and islet cells. (C) Cipralex plus SIL group: demonstrating moderate immunoreactivity in pancreatic acini and islet cells as compared with cipralex group (×400, scale bar=20 μm). (D) A histogram illustrating a highly significant increase in caspase-3 immune expression in cipralex group compared to control group. While cipralex plus SIL group demonstrating a highly significant decrease compared to cipralex group, with no significance between control and cipralex plus SIL groups. SIL, silymarin.

• 

  Fig. 11 Representative PCNA immunostaining of rat pancreas; (A) Control group: illustrating strong nuclear immunoreactivity pancreatic acini and islet cells. (B) Cipralex group: showing mild immunoreactivity for PCNA in most of pancreatic acini and islet cells. (C) Cipralex plus SIL group: showing moderate immunoreactivity pancreatic acini and islet cells as compared with cipralex group (×400, scale bar=20 μm). (D) A histogram exhibiting a highly significant decrease in PCNA immune expression in cipralex group compared to control group. While cipralex plus SIL group showing a highly significant increase compared to cipralex group, with no significant difference between control and cipralex plus SIL groups. PCNA, proliferating cell nuclear antigen; SIL, silymarin.

• 

  Fig. 12 Representative insulin immunostaining of rat pancreas; (A) Control group: revealing strong cytoplasmic immunoreactivity for insulin in most of the cells of the pancreatic islet. (B) Cipralex group: showing mild immunoreactivity for insulin in a small number of islets cells. (C) Cipralex plus SIL group: showing moderate cytoplasmic immunoreactivity for insulin in most of the pancreatic islet as compared with cipralex group (×400, scale bar=20 μm). (D) A histogram exhibiting a highly significant decrease in insulin positive β-cells immune expression in cipralex group compared to control group. While cipralex plus SIL group showing a significant increase compared to cipralex group, with no significant difference between control and cipralex plus SIL groups. SIL, silymarin.

• 

  Fig. 13 A histogram showing a highly significant decrease (P<0.001) in the number and diameter of islets of Langerhans in the cipralex group compared to the control group, while the cipralex plus SIL group, illustrating a significant increase compared to the cipralex group, with no significant difference between control and cipralex plus SIL groups. SIL, silymarin.

Reference

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